Abstract: Objective To establish the tetracycline and its derivatives-controlled inducible gene expression system in human pancreatic cancer line. Methods A eukaryotic expression vector for highly expressed tranactivtor was constructed and transfected into PANC-1 cell line by Fugene HD.Transfected cells were selected in the medium containing 2μg/ml puromycin and isolated after 2 weeks.All individual clones were screened with the luciferase reporter gene system for low-background and highly controlled clones.Then,tTA expression in these clones was detected by RT-PCR and cell immunofluorescence,respectively. Results Three doxcycline-controlled individual PANC-1 cell lines with high tTA expression were isolated and identified by luciferase reporter gene assay.RT-PCR and cell immunofluorescence showed the tTA expression in the 3 clones. Conclusion Human pancreatic cancer lines with low background and highly controlled clones are successfully established,thus providing an effective way for gene function studies.